The leading cause of death in diabetic patients is cardiovascular disease (CVD). Our long-term goal is to identify new pharmacological and dietary strategies for the prevention of CVD in diabetic patients. LDL receptors play a central role in cholesterol homeostasis. Using liver insulin receptor knockout mice, we previously showed that insulin, in the presence of a high-fat, high-cholesterol diet, produces a five-fold increase in LDL receptor protein which correlates with improved LDL clearance and protection from atherosclerosis (Biddinger et al., Cell Metab., 2008). That insulin can exert such a profound, protective effect upon LDL receptor protein was unexpected. Mylip is an E3 ubiquitin ligase that has recently emerged from GWAS studies as an important regulator of lipoprotein metabolism and found to ubiquitinate and degrade the LDL receptor. The overall goal of this proposal is to determine the role of Mylip in producing the dyslipidemia and atherosclerosis associated with the diabetic state. We present strong preliminary data that insulin suppresses Mylip mRNA in multiple in vitro and in vivo models and that diabetic mice show increased Mylip expression, reduced LDL receptor protein and hypercholesterolemia. Based on this, we hypothesize that insulin suppresses Mylip transcription and that increased Mylip in the diabetic state leads to decreased LDL receptor protein, dyslipidemia and atherosclerosis. Our aims are as follows: (1) use luciferase assays to determine the mechanisms by which insulin regulates Mylip transcription; (2) determine the role of Mylip in mediating the effects of insulin on LDL receptor ubiquitination and half-life in vitro; and (3) determine the extent to which knockdown of Mylip in diabetic mice can prevent dyslipidemia and atherosclerosis. Mylip potentially represents a key regulatory node between insulin and the LDL receptor that could be exploited for therapeutic intervention. The proposed studies will provide important insights into the regulation of Mylip and its role in the diabetic state.